recent patents on anti-cancer drug discovery, 2019, 14, 5-18 · tts are wheat germ, grapefruit,...

Re

cent

Pat

ents

on

Anti-

Can

cer

Dru

g D

isco

very

��%��;>?=5@AB@��%��BB;B5<A?C

��

Send Orders for Reprints to [email protected]

Recent Patents on Anti-Cancer Drug Discovery, 2019, 14, 5-18

5

REVIEW ARTICLE

Tocotrienols and Cancer: From the State of the Art to Promising Novel

Patents

Fabrizio Fontana, Michela Raimondi, Monica Marzagalli, Roberta M. Moretti, Marina Montagnani Marelli and Patrizia Limonta*

Department of Excellence, Department of Pharmacological and Biomolecular Sciences, University of Milan, Milan, Italy

A R T I C L E H I S T O R Y

Received: October 31, 2018 Revised: January 04, 2019 Accepted: January 04, 2019 DOI: 10.2174/1574892814666190116111827

Abstract: Background: Tocotrienols (TTs) are vitamin E derivatives naturally occurring in several plants and vegetable oils. Like Tocopherols (TPs), they comprise four isoforms, α, β, γ and δ, but unlike TPs, they present an unsaturated isoprenoid chain. Recent studies indicate that TTs provide important health benefits, including neuroprotective, anti-inflammatory, anti-oxidant, cholesterol lowering and im-munomodulatory effects. Moreover, they have been found to possess unique anti-cancer properties.

Objective: The purpose of this review is to present an overview of the state of the art of TTs role in cancer prevention and treatment, as well as to describe recent patents proposing new methods for TTs isolation, chemical modification and use in cancer prevention and/or therapy.

Methods: Recent literature and patents focusing on TTs anti-cancer applications have been identified and reviewed, with special regard to their scientific impact and novelty.

Results: TTs have demonstrated significant anti-cancer activity in multiple tumor types, both in vitro and in vivo. Furthermore, they have shown synergistic effects when given in combination with stan-dard anti-cancer agents or other anti-tumor natural compounds. Finally, new purification processes and transgenic sources have been designed in order to improve TTs production, and novel TTs formula-tions and synthetic derivatives have been developed to enhance their solubility and bioavailability.

Conclusion: The promising anti-cancer effects shown by TTs in several preclinical studies may open new opportunities for therapeutic interventions in different tumors. Thus, clinical trials aimed at con-firming TTs chemopreventive and tumor-suppressing activity, particularly in combination with stan-dard therapies, are urgently needed.

Keywords: Cancer prevention, cancer therapy, natural compounds, tocotrienols, tocotrienol formulations, tocotrienol synthetic derivatives.

1. INTRODUCTION

The term “nutraceutical” was coined in 1989 by Dr. Ste-phen De Felice, MD founder and chairman of the Foundation for Innovation in Medicine, who defined it as a “food, or parts of food, that provide medical or health benefits, includ-ing the prevention and treatment of disease”. Thus, nutraceu-ticals may refer to naturally nutrient-rich or biologically active foods and herbs, such as blueberries or soybeans, or they may be specific food components or isolated nutrients and phytochemicals with medicinal properties, such as ω-3 fatty acids, polyphenols, flavonoids, carotenoids and vita-mins [1].

*Address correspondence to this author at the Department of Excellence, Department of Pharmacological and Biomolecular Sciences, University of Milan, Milan, Italy; Tel: +390250318213; Fax: +390250318204; E-mail: [email protected]

Vitamin E family consists of eight members, α-, β-, γ- and δ-Tocopherols (TPs) and the corresponding four Tocotrienols (TTs). TTs, in particular, were demonstrated to possess spe-cific health-promoting and/or disease-preventing properties in different pathologies, associated with their powerful neuropro-tective, anti-inflammatory, anti-oxidant, cholesterol lowering and immunomodulatory potentials [2]. It has also been ob-served that TTs, particularly γ and δ isoforms, can exert a po-tent anti-cancer activity in different tumors [3]. The molecular mechanisms of these anti-proliferative properties remain to be fully elucidated and they appear to involve several intracellu-lar pathways [4, 5] (Fig. 1).

This review aims to describe the anti-tumor effects of TTs based on the experimental evidence so far available, as well as to offer a selection of published patents in the last years, which provide new methods for the production and use of TTs in cancer prevention and/or treatment.

2212-3970/19 $58.00+.00 © 2019 Bentham Science Publishers

http://crossmark.crossref.org/dialog/?doi=10.2174/1574892814666190116111827&domain=pdf

6 Recent Patents on Anti-Cancer Drug Discovery, 2019, Vol. 14, No. 1 Fontana et al.

2. TOCOTRIENOLS

Vitamin E was discovered by Herbert Evans and Katharine Bishop in 1922 [6] and isolated by Evans and Gladys Emerson in 1935 [7]. Its hydrophobic components are divided into two groups: α-, β-, γ- and δ-TPs and α-, β-, γ- and δ-TTs. The critical chemical structural difference be-tween them is that the chromanol ring, common to both groups, is linked to a saturated isoprenoid side chain in TPs and to an unsaturated isoprenoid side chain in TTs. Moreo-ver, each isoform of both TPs and TTs differs from others in the number and position of methyl groups on the chromanol ring: the α and β isomers are trimethylated, the γ isomers are dimethylated and the δ isomers are monomethylated.

The metabolism of TTs is still not clear. They are ab-sorbed in the small intestine in the presence of bile salts and transported to body tissues through the blood after α -‐TP transport protein-mediated sequestration into liver lipopro-teins. However, TTs seem to have lower affinity than TPs for α-‐TP transport protein, and various studies have pointed out that TPs can interfere with TTs benefits by lowering their intestinal absorption and increasing their catabolism in the liver [8, 9]. Despite these observations, preclinical trials have reported high activity of these compounds after oral admini-stration, without significant side effects and toxicity [10, 11]. High bioavailability and safety of TTs were also demon-strated in healthy human subjects and patients with breast and pancreatic cancer [12-14].

In nature, TTs are present in many plants, cereals, seeds, nuts and grains, as well as in the oils derived from them.

They are particularly abundant in palm oil, which represents the main source of TTs and in particular of γ -TT [15], an-natto (Bixa orellana L.) seeds, which contain about 150mg δ-‐TT/100g dry seeds with no TPs [16], and rice bran, con-taining high levels of α- and γ-TT [17, 18]. Other sources of TTs are wheat germ, grapefruit, hazelnuts, olive oil, sun-flower oil and flaxseed oil [19, 20].

2.1. Tocotrienols in Cancer

In recent years, TTs have emerged as one of the most ef-fective class of natural compounds for preventing and ame-liorating cardiovascular and neurodegenerative diseases, as well as hyperlipidemia, inflammation, diabetes and osteopo-rosis [21-24]. In addition, they have demonstrated promising potential as anti-cancer agents, particularly in breast, cervix, colon, gastric, lung, skin, pancreatic and prostate cancer.

2.1.1. Tocotrienols in Breast Cancer

γ-‐TT was found to exert both anti-proliferative and pro-apoptotic activity in breast cancer cells. In particular, it was shown to reduce the levels of cyclin D1 and Cy-clin-‐Dependent Kinases (CDK) 2, 4 and 6 and to increase the expression of CDK inhibitors [25, 26]; to suppress both the PI3K/Akt/mTOR and the Ras/Raf/MEK/ERK signaling pathways and to decrease c-‐Myc levels [27]; to induce intrinsic apoptosis accompanied by cytochrome c release, mitochondrial membrane depolarization, caspase activation, DNA fragmentation and poly(ADP-ribose) polymerase cleavage [28, 29]; to trigger the extrinsic apoptotic pathway by activation of caspase-8 [30].

Fig. (1). Molecular mechanisms of Tocotrienols (TTs) anti-cancer activity.

��

�� ! �� "# �� "$ �� "%�$! ��$&

��

��

��

'��( �)*�% �)*�� )(�!�+'��,��-��.��

��

�� /)�0�! �/)�0�$��0�$ ��0��/�.��1��

��

�2��.��1��

Tocotrienols and Cancer Recent Patents on Anti-Cancer Drug Discovery, 2019, Vol. 14, No. 1 7

δ-‐TT was also demonstrated to exert potent anti-tumor effects in mammary cancer cells, by reducing proliferation through downregulation of the HMG-‐CoA reductase activity and inhibition of cholesterol synthesis [31] and by inducing oxidative stress-related mitochondrial apoptosis [32].

The Human Epidermal Growth Factor Receptor 2 (HER-2) is a receptor tyrosine-protein kinase that in humans is en-coded by the erbB-2 gene and normally promotes breast cell proliferation. Amplification of this oncogene occurs in 30% of breast tumors, thus representing an important biomarker and target for therapy [33]. Alawin et al. demonstrated that HER-2 receptors and TTs specifically accumulate in breast cancer cell lipid raft microdomains. Moreover, they found that TTs profoundly alter the composition of the lipid rafts, with subsequent disruption of their integrity and inactivation (due to reduced dimerization and phosphorylation) of the associated HER-2 receptors and of the downstream signaling pathways [34].

Almost 70% of human breast cancers are estrogen-dependent and estrogen receptor-positive. TTs were shown to promote the nuclear translocation of the anti-proliferative Estrogen Receptor (ER) β and to decrease the tumorigenic ERα expression [35]. On the other hand, Khallouki et al. suggested that δ-‐TT can induce cytotoxic effects in breast cancer cells independently of their ER status [31].

A proper protein folding and processing are essential for the maintenance of cellular homeostasis, and two important mechanisms involved in its control are the Unfolded Protein Response (UPR), activated in case of Endoplasmic Reticu-lum (ER) stress, and autophagy. The ER stress is a cellular process that can be triggered by various physiological condi-tions and pathological insults, as well as by different syn-thetic and natural drugs: Cells initially respond with a defen-sive UPR, accompanied by an increase in the ER protein folding capacity and in the ER-Associated Protein Degrada-tion (ERAD) machinery; however, in case of severe or pro-longed stress, unfolded and misfolded proteins may exceed the ER capacity and accumulate in its lumen, leading to the activation of a set of pro-death programs, such as the double-stranded RNA-dependent Protein Kinase PKR-Like ER Kinase (PERK)/Eukaryotic Initiation Factor 2α (eIF2α)/Activating Transcription Factor 4 (ATF4)/C/EBP homologous protein (CHOP) pathway and the Inositol-Requiring Enzyme 1α (IRE1α)/c-‐Jun N-‐Terminal Kinase (JNK)/p38 MAPK cascade [36]. Autophagy is an evolution-arily conserved catabolic process that is used to deliver cyto-plasmic material, such as damaged organelles and protein aggregates, to the lysosome for degradation: it is character-ized by the formation of double-membrane vesicles, the autophagosomes, that fuse with lysosomes for cytoplasmic cargo recycling, and it is mediated by two proteins, microtu-bule-associated proteins 1A/1B light chain 3B, commonly known as LC3, and p62 (also called SQSTM1) [37]. Phar-macological targeting of these two molecular pathways has been recently proposed as an effective treatment strategy for tumors [38, 39]. Different natural compounds were reported to induce ER stress- and autophagy-mediated death in cancer cells [40, 41]: among them, γ-‐TT specifically triggered both of these pro-apoptotic pathways in breast cancer cells [42-46].

Angiogenesis, the process that leads to new blood vessel formation from pre-existing vasculature, and metastatization, the spread of cancer cells from the tissue of origin to new areas of the body, are two main steps in tumor growth and progression. In breast cancer, TTs exhibited both anti-metastatic and anti-angiogenic properties, associated with the inhibition of Met/hepatocyte growth factor receptor and Rac1/WAVE2 cascade and the downregulation of VEGF expression, respectively [47-50].

It is known that Cancer Stem Cells (CSCs) represent only a small subpopulation of cancer cells within a tumor mass. However, their self-renewal ability and their capacity to differentiate into the entire heterogeneous tumor cell bulk make them important therapeutic targets. Moreover, they appear to be implicated in the resistance to standard cancer treatments, also promoting tumor recurrence and metastasis [51]. Many natural compounds were found to target CSCs [52], and TTs were shown to specifically eliminate the breast CSCs subpopulation, alone or in combination with simvas-tatin [53, 54].

In +SA mammary tumor cells TTs treatment synergisti-cally increased the anti-cancer activity of synthetic drugs, such as tyrosine kinase inhibitors (erlotinib and gefitinib), statins (simvastatin, mevastatin and lovastatin) and the COX-2 selective nonsteroidal anti-inflammatory agent celecoxib, through suppression of HERB2-4 receptors expression levels and inhibition of Akt and MAPK pathways [55-57]. Fur-thermore, in multidrug-resistant MCF-7/ADR cells γ-TT significantly reduced the expression of P-gp, leading to en-hanced accumulation of doxorubicin in cells and subsequent G2/M cell cycle arrest and apoptosis [58]. Similar synergistic effects were shown by TTs when given in combination with several natural compounds: in murine malignant mammary epithelial cells, the co-treatment with sesamin, a lignan con-tained in sesame seeds and oil, not only improved TTs bioavailability by reducing their metabolic degradation but also exhibited a synergistic inhibitory effect on the EGF-dependent proliferative pathway [59-61]; the addition of a polyphenol, such as Epigallocatechin Gallate (EGCG), found in green tea, or resveratrol, present in berries and grapes, potentiated the γ-TT-induced downregulation of cyclin D1 and Bcl-2 expression in MCF-7 human breast cancer cells, and the triple combination of these compounds synergisti-cally up-regulated the expression of NAD(P)H Quinone De-hydrogenase 1 (NQO1), an enzyme activated in case of re-dox imbalance [62]; a combination of TTs and oridonin caused a significant additive effect in decreasing +SA cell viability through suppression of Akt/mTOR signaling and elevation of apoptosis (caspase-3 and Bax/Bcl-2 ratio) and autophagy (Atg and Beclin-1) markers [63].

It should be noted that also semisynthetic redox-silent TT oxazine derivatives were found to successfully inhibit breast tumor growth, both in vitro and in vivo [64-66]. In particular, they successfully counteracted the CoCl2-induced increase of HIF-‐1α levels, with a parallel inactivation of the Akt/mTOR pathway and of its downstream targets p70S6K and eIF-4E1. In addition, TT oxazine derivative treatment resulted in a blockade of the CoCl2-mediated VEGF overex-pression.


A five-year double-‐blinded and placebo-‐controlled clini-cal trial was conducted in 250 women with early breast can-cer to investigate the TTs adjuvant potentials when given in combination with tamoxifen [67]. The patients, 40-60 years old, with either stage I or II estrogen receptor-positive breast cancer, were non-randomly assigned to two groups: The treatment group was administered 400mg/day TRF plus ta-moxifen while the control group was given placebo plus ta-moxifen. The 5-‐year breast cancer-‐specific survival was 98.3% in the treatment group and 95% in the control group, while the 5-‐year disease-‐free survival was 86.7% and 83.3%, respectively. The mortality risk was 60% lower in the TRF group versus controls but it was not statistically significant, probably as a result of the small sample size of the experi-ment.

2.1.2. Tocotrienols in Cervical Cancer

TTs were reported to inhibit HeLa cervical cancer cell proliferation through downregulation of the expression of cell cycle-related proteins, such as cyclin D3, p16 and CDK6. Moreover, the induction of HeLa cell death by TTs appeared to be associated with the upregulation of Interleu-kin-6 (IL-6) [68].

Comitato et al. demonstrated that γ- and δ-TT induce apoptosis in cervical cancer cells, by triggering molecular signals associated with ER stress, such as IRE-1α phos-phorylation, XBP-1 alternative splicing and CHOP enhanced transcription [69]. Furthermore, they observed significant Ca2+ release from the ER membranes to the cytoplasm, as well as an interesting modulation of isoprenoid, sterol and steroid biosynthesis and metabolism, with SCD, LPIN and SREBPF1-2 downregulation.

γ-‐TT was found to specifically target Src Homology 2 Domain-Containing Phosphatase 2 (SHP2) and the RAS/ERK signaling pathway in spheres from cervical can-cer, inhibiting the CSCs subpopulation growth [70].

2.1.3. Tocotrienols in Colon Cancer

In RKO human colon cancer cells, a TRF preparation triggered intrinsic apoptosis correlated with p53 and caspases activation, Bax/Bcl-‐2 ratio modulation, chromatin condensa-tion, DNA fragmentation and cell membrane shrinkage [71]. Moreover, γ-TT was reported to profoundly alter sphin-golipid metabolism in HCT-116 cells through suppression of dihydroceramide desaturase activity and activation of sphin-gomyelin hydrolysis, ultimately leading to autophagy and apoptosis [72].

Paraptosis is a type of programmed cell death character-ized by cytoplasmic vacuolization, particularly by ER dilata-tion and mitochondrial swelling. Various studies have shown that several natural compounds, such as celastrol, paclitaxel, ophiobolin A and curcumin, can induce this non-apoptotic cell death in different types of tumors [73]. It has been re-cently demonstrated that also TTs can trigger paraptosis in SW620 and HCT-8 human colon carcinoma cells through inhibition of the Wnt signaling pathway and downregulation of c-jun, cyclin D1 and β-catenin levels [74, 75].

A synergistic antitumor activity of γ-‐TT and different synthetic and natural anti-cancer agents was observed: the

co-treatment with capecitabine synergistically decreased Ki-67, cyclin D1, NF-κB, CXCR4 and MMP-9 expression lev-els in a nude mouse xenograft model of human colorectal cancer [76]; addition of atorvastatin to tocotrienol treatment enhanced the disruption of RhoA signal transduction in HT29 and HCT116 human colon cancer cells, and a triple combination with celecoxib resulted in a synergistic induc-tion of G(0)/G(1) phase cell cycle arrest and apoptosis [77]; 6-gingerol, the bioactive constituent of ginger, potentiated the γ-‐TT pro-apoptotic activity in HT29 and SW837 human colorectal cancer cell lines, inducing caspase-3 activation and significant morphological changes, such as cell shrink-age and pyknosis [78].

δ-‐TT was shown to suppress hypoxia-induced VEGF, IL-8 and COX-2 synthesis in DLD-1 human colorectal ade-nocarcinoma cells [79], as well as to inhibit tube formation, migration and adhesion of HUVEC cells grown in DLD-1 conditioned medium [80]. These results were also confirmed by in vivo experiments [80, 81].

Rice bran is not only enriched in δ-TT but also in ferulic acid. Eitsuka et al. demonstrated that co-treatment with both these natural compounds significantly increased the intracel-lular concentration and anti-tumor activity of δ-TT in DLD-1 cells, suggesting that ferulic acid improves the bioavailabil-ity as well as the therapeutic effectiveness of δ-TT [82]. In particular, this combination treatment was shown to syner-gistically down-regulate the expression of Human Telom-erase Reverse Transcriptase (hTERT), the catalytic subunit of telomerase, thus suppressing its proliferative activity.

2.1.4. Tocotrienols in Gastric Cancer

γ-‐TT was first demonstrated to induce intrinsic apoptosis in human gastric cancer cells through the suppression of the MAPK signaling [83, 84].

γ-‐TT was also found to exert potent anti-metastatic and anti-angiogenic activity in gastric cancer. In particular, it in-hibited cell migration and invasion capability, by reducing the expression of the matrix metalloproteinases MMP-‐2 and MMP-‐9 and by increasing the levels of Tissue Inhibitor of Metalloproteinase-‐1 (TIMP-‐1) and TIMP-‐2 [85], and signifi-cantly counteracted the hypoxia-mediated HIF-‐1α over-expression and VEGF synthesis by modulation of the ERK signaling pathway [86]. Furthermore, it significantly de-creased the expression of VEGFR-2 in HUVEC cells grown in a conditioned medium of gastric adenocarcinoma cells [87].

In addition to its pro-apoptotic, anti-metastatic and anti-angiogenic effects, γ-‐TT was reported to enhance the antitu-mor activity of capecitabine in human gastric cancer cell lines, as well as in nude mice xenografted with human gas-tric cancer cells [88].

2.1.5. Tocotrienols in Lung Cancer

MicroRNAs (miRNAs) are endogenous, ~22 nucleotides, non-coding RNAs that play key regulatory roles in animals and plants by inducing transcriptional silencing through mRNAs cleavage or translational arrest. miRNAs may func-tion as either oncogenes or tumor suppressors (oncomirs), depending on the specific cancer type [89]. Ji et al. observed that δ-‐TT could inhibit nonsmall cancer cell growth and in-


vasiveness through upregulation of miR-34a, which resulted in decreased expression of Notch-1 and its downstream tar-gets, such as Hes-1, cyclin D1, survivin and Bcl-2 [90]. Moreover, they found that δ-‐TT exhibited a significant syn-ergistic anti-cancer effect when given in combination with cisplatin. This was related to a reduction of the NF-κB DNA binding activity and to an increase in cleaved caspase-3 and PARP expressions [91].

TTs were also shown to potentiate lovastatin-mediated cell growth arrest in the A549 human lung carcinoma cell line [92].

2.1.6. Tocotrienols in Pancreatic Cancer

Anti-proliferative and pro-apoptotic effects of δ-‐TT, me-diated by p27Kip1-dependent cell cycle arrest [93], inhibition of HMG-CoA reductase activity [94] or HER2 [95] and EGR-1/Bax pathway activation [96], were shown in pancre-atic cancer cells. Similar results were obtained after γ-‐TT treatment [97].

TTs significantly suppressed the invasive behavior through downregulation of specific epithelial-mesenchymal transition (EMT) biomarkers (such as N-cadherin, vimentin and MMP9) in L3.6pl and Mia PaCa-2 cells, both in vitro and in vivo [98].

δ-‐TT successfully targeted and eliminated Pancreatic Duc-tal Adenocarcinoma (PDAC) stem-‐like cells, decreasing the expression of CSCs self-renewal-promoting transcription fac-tors, Oct4 and Sox2, and delaying tumor onset in mice [98].

Promising results were reported in pancreatic cancer cells treated with polyethylene glycol (350 and 1000) succi-nate derivatives of TTs [99]. In addition, in vitro studies pointed out that entrapment of gemcitabine/γ-tocotrienol or paclitaxel/γ-tocotrienol lipid conjugates into nanoemulsions significantly enhanced their anti-tumor effects when com-pared to the free drug [100, 101].

Springett et al. investigated δ-‐TT efficacy and safety in patients with PDAC [14]. In this Phase I preoperative clinical trial, 25 patients were given crescent oral doses of δ-‐TT (from 200 to 3200mg) daily for 13 days before surgery and one dose on the day of surgery. Except for one case of drug-related grade 1 diarrhea registered at the higher daily dose level, the treatment was well tolerated, with no dose-limiting toxicity. δ-‐TT exhibited an effective half-life of about 4 hours, rapidly reaching bioactive levels in blood and inducing apoptosis-associated caspase-3 cleavage in cancer cells. In particular, the most effective δ-‐TT dose was between 400 and 1600mg.

2.1.7. Tocotrienols in Prostate Cancer

TTs were reported to exert anti-proliferative and pro-apoptotic effects in different prostate cancer cell lines by targeting several signaling pathways, such as Phosphoinosi-tide-‐3 Kinase (PI3K)/Akt/mTOR, Signal Transducer and Activator of Transcription (STAT), transforming growth factor β (TFGβ) receptor and NF-‐κB cascades, as well as cyclins and the cell cycle inhibitors p27 and p21 [102-106].

It has been shown that γ-‐TT significantly decreases the expression of CD133 and CD44 CSCs markers in PC-3 and DU145 castration-‐resistant prostate cancer cells, also sup-

pressing their anchorage-independent growth and spheroidogenic ability. Moreover, γ-‐TT pre-treatment of pros-tate cancer cells resulted in the inhibition of tumor initiation after their inoculation in nude mice. Finally, despite the high resistance of CD133-‐positive PC-‐3 cells to docetaxel treat-ment, they were as sensitive to γ-‐TT as the CD133-‐depleted population [107]. Similar studies were performed by Lee et al., who confirmed the γ-‐TT ability to specifically target the CSCs subpopulation in different prostate cancer cell lines and mouse models, leading to a significant suppression of the pro-liferation of the castration-‐resistant tumors [108].

Recent evidence suggests that also δ-‐TT can inhibit the proliferation of prostate CSCs under hypoxic conditions, by specifically inactivating the HIF-1α cascade [109].

In prostate cancer, TTs were reported to potentiate the anti-tumor activity of lovastatin in vitro [110]. A combination treatment with δ-‐TT and geranylgeraniol was also shown to synergistically suppress the viability of DU145 prostate cancer cells, inducing a significant downregulation of the expression of HMG-CoA reductase, as well as an interesting reduction in membrane K-RAS protein levels [111].

2.1.8. Tocotrienols in Skin Cancer

We recently demonstrated that δ-‐TT induces ER-stress-mediated apoptosis in human melanoma cells in vitro and in tumors in vivo, through the activation of the PERK/p-eIF2α/ATF4/CHOP, IRE1α and caspase-4 ER stress-related branches [112]. Moreover, we observed that, unlike vemu-rafenib, it can selectively target and eliminate the melanoma ABCG2-positive CSCs subpopulation, successfully inducing disaggregation of A375 melanospheres and reducing the spheroid formation ability of sphere-derived cells [113].

γ-‐TT also exhibited apoptosis-inducing and invasion-suppressing activity in malignant melanoma cells. In particu-lar, it was reported to inhibit NF-κB, EGF-R and Id family proteins, to activate the JNK signaling pathway, to decrease different mesenchymal markers levels and to restore E-cadherin and γ-catenin expression [114].

The Aryl hydrocarbon Receptor (AhR) is a ligand-activated transcription factor, which controls many biologi-cal and physiological processes in response to aromatic hy-drocarbons, such as cellular proliferation and differentiation, tissue development, immune and toxic response and skin barrier homeostasis. Upon ligand binding to AhR, the acti-vated complex translocates to the cell nucleus, where it forms a functional heterodimer with the AhR nuclear trans-locator (ARNT), with subsequent interaction with DNA and transcriptional activation of several target genes (in particu-lar p21 and Bax) by binding to the xenobiotic responsive element [115]. Yamashita et al. demonstrated that γ-‐TT in-duces AhR expression in a dose-dependent manner in B16 mouse melanoma cells and enhances its sensitivity to baical-ein, a flavone particularly abundant in the roots of Scutel-laria baicalensis and Scutellaria lateriflora, which can in-hibit tumor cell growth by acting as a ligand of AhR and thus upregulating p21 and Bax levels [116].

Synergistic antitumor activity of TTs and lovastatin was evidenced in murine B16 melanoma cells, as well as in C57BL6 mice bearing B16 xenografts [92].


Improved anti-proliferative effectiveness against A431 and SCC-4 human keratinocyte cancer cells in vitro was shown by a hybrid-nanoemulsified TTs delivery system and it was associated with better yield in physicochemical parame-ters, as well as stability in chemical and structural composition [117]. Interestingly, an orally administered γ-TT nano-formulation also exhibited enhanced radioprotection compared to γ-TT alone in CD2F1 mice exposed to total body γ radiation [118].

Transferrin receptors are frequently expressed in tumor cells, thus representing a potential target for the delivery of anti-cancer drugs into the tumor mass. Karim et al. reported that α-‐TT entrapped in transferrin-‐bearing multilamellar vesicles possesses potent growth-suppressing activity in A431 human epidermoid carcinoma cancer cells and B16-F10 murine melanoma cells. Moreover, the intravenous ad-ministration of these vesicles to mice bearing A431 and B16-F10 tumors successfully inhibited cancer progression, with-out visible side effects [119].

2.1.9. Tocotrienols in Other Tumors

Anti-proliferative, pro-apoptotic, anti-metastatic and anti-angiogenic effects of TTs were also reported in other cancer cell types.

In bladder cancer cells, δ-TT treatment suppressed cell proliferation and colony formation capacity by triggering G1 phase arrest and intrinsic apoptosis, accompanied by upregu-lation of p21, p27 and Bax expression and downregulation of cyclin D1, Bcl-2, Bcl-xL and Mcl-1 levels, ultimately lead-ing to caspase-3 and PARP cleavage. Moreover, δ-TT in-

duced ETK inactivation and SHP-1 increase, resulting in the inhibition of the STAT3 pathway. Importantly, low doses of δ-TT sensitized human bladder cancer cells to the anti-tumor activity of gemcitabine [120].

Tan et al. demonstrated that γ-TT can act as a BH3 mi-metic in SH-SY5Y neuroblastoma cells, by binding to the hydrophobic groove of Bcl-2 and triggering Bax- and caspase-9-mediated apoptosis [121].

TTs were shown to suppress cell viability in different leukemic cell lines and blasts from patients, by inducing loss of mitochondrial membrane potential, release of cytochrome C and cleavage of Bid and caspases. Interestingly, normal mononuclear cells were spared by TTs-induced cytotoxic effects [122].

γ-TT triggered apoptotic cell death in human T-‐cell lym-phoma via both the intrinsic and extrinsic pathways. In par-ticular, the tocotrienol treatment resulted in mitochondrial ROS production, calcium cytoplasmic accumulation, JNK phosphorylation and changes in the Bax/Bcl-‐2 ratio, as well as in Fas and FasL upregulation and caspase-‐8 activation [123].

In human hepatocellular carcinoma cells, TTs exerted potent anti-proliferative effects correlated with a significant decrease of the expression levels of H-RAS, Grb2, Sos-1, p-Src and p-Shc and subsequent inhibition of the RAS-RAF-MEK-ERK downstream signaling pathway [124]. Moreover, γ-TT significantly reduced the angiogenesis-mediated growth of human hepatocellular carcinoma in an orthotopic mouse model by suppressing the AKT/mTOR [125] pathway (Table 1) [65-125].

Table 1. In Vivo and Clinical Studies on Tocotrienols (TTs) in Cancer Therapy/Prevention.

Tumor Type Current Studies References

Breast cancer

• Two in vivo studies demonstrating the anti-proliferative and anti-angiogenic effects of redox-silent TT oxazine derivatives

• One clinical trial aimed at investigating the TTs adjuvant potentials when given in combination with tamoxifen

[65-67]

Colon cancer • One in vivo study showing the synergistic anti-cancer effects of TTs and capecitabine

• Two in vivo studies demonstrating the anti-angiogenic properties of TTs [76, 80, 81]

Gastric cancer • One in vivo study showing the synergistic anti-cancer effects of TTs and capecitabine [88]

Lung cancer • One in vivo study showing the synergistic anti-cancer effects of TTs and lovastatin [92]

Pancreatic cancer

• One in vivo study demonstrating the pro-apoptotic properties of TTs

• One in vivo study demonstrating the anti-invasive properties of TTs

• One clinical trial conducted to test TTs efficacy and safety

[14, 97, 98]

Prostate cancer • Two in vivo studies demonstrating the anti-proliferative properties of TTs

• Two in vivo studies demonstrating the specific CSCs-targeting ability of TTs [103, 106-108]

Skin cancer

• One in vivo study demonstrating the pro-apoptotic properties of TTs

• One in vivo study demonstrating the specific CSCs-targeting ability of TTs

• One in vivo study demonstrating the chemopreventive properties of TTs

• One in vivo study demonstrating the anti-cancer effects of transferrin-bearing vesicles containing TTs

[112, 113, 118, 119]

Liver cancer • One in vivo study demonstrating the anti-proliferative and anti-angiogenic effects of TTs [125]


3. PATENTS FOR THE ISOLATION, CHEMICAL MODIFICATION AND USE OF TOCOTRIENOLS IN CANCER PREVENTION/THERAPY

In the last years, novel methods and techniques have been proposed to successfully isolate and exploit TTs in cancer prevention and therapy.

3.1. Isolation and Purification of Tocotrienols

As reported above, TTs are present in several plants and vegetable oils, and many studies have highlighted the need for development of effective isolation and purification proc-esses for the proper preparation of TTs intended for experi-mental or clinical testing [20, 126, 127].

Red palm oil is the major source of TTs among all edible oils, with TTs representing 70% of total vitamin E derivatives present in it. Different patents provide a method for the extrac-tion of TTs from Palm Fatty Acid Distillates (PFAD), which involves conversion of a PFAD into distilled methyl esters, followed by ion-exchange chromatography and by another step of molecular distillation to get vitamin E components concentrate of more than 90% [128-134]. Some other patents propose to obtain TTs from Crude Palm Oil (CPO) by con-verting CPO to methyl esters, with subsequent removal of the esters by distillation to yield carotene-rich and vitamin E-rich fraction [135-137]. Conversion of CPO into methyl esters, followed by chromatographic separation in the presence of solvents in order to obtain carotene-rich, vitamin E-rich and sterol-rich fractions, has also been suggested [138]. Finally, two novel inventions disclose the use of supercritical fluids, such as SC-CO2, in the adsorption/desorption chromatography isolation/separation of TTs from CPO [139, 140].

High levels of TTs can be easily obtained from rice bran through its ultrasound-assisted saponification and subsequent recrystallization or chromatography separation of the saponi-fied organic phase content, as shown in some new patents [141, 142]. This process also offers the possibility to suc-cessfully extract ferulic acid from the aqueous layer obtained in the above separation process, by precipitation of its alkali salt through addition of an acid, such as diluted sulfuric or hydrochloric acid [142].

TTs isolation from annatto seeds has been described in a recent invention [143]. This process presents many advan-tages. First of all, the amount of δ-‐TT present in the byprod-uct solution of Bixa Orellana L. seeds is higher than that found in other common sources, such as palm or rice bran oil. Moreover, and also in contrast to palm and rice bran oils, Bixa Orellana L. seeds extract essentially does not contain α-‐TP, which can have a mitigating effect on the therapeutic properties of δ-‐TT, as discussed above. Finally, the annatto solution is also a convenient source of geranylgeraniol, that can be administered in combination with TTs in order to potentiate their anti-tumor effects.

A novel patent highlights a method for producing a tocotrienol-rich composition by transesterification of coconut palm oil, followed by distillation of the reaction product and adsorption treatment of the residue with an anion exchange resin having a 2-hydroxyethyl dimethylammonium group [144].

A new invention provides a procedure for the production of purified γ-‐ and/or δ-‐TT from TTs rich oils and distillates [145]. In this process, γ-‐ and/or δ-‐TT are rapidly isolated

from the TTs sources by medium (flash) or low-pressure chromatography, obtaining a fraction with high concentra-tion of γ-TT and/or δ-TT and minimized content of α-isomers. In particular, the γ- and/or δ-TT rich composition contains approximately 95% of TTs, with 10% of γ-TT and 3% of δ-TT, each of which having purity of about 95%.

3.2. Transgenic Sources of Tocotrienols

The use of biotechnological techniques to increase the bioactive components content in plants has become a very popular subject of study and debate in the last decades [146].

A recent patent reports on a method for the production of a transgenic rice plant enriched in δ-TT, whose extract suc-cessfully inhibits PC-3 prostate cancer cell growth. The transformed plant is obtained through the introduction in its DNA of the Homogentisic Acid Geranylgeranyl Transferase (HGGT) gene, which mediates TTs synthesis by catalyzing the condensation of homogentisate and geranylgeranyl diphosphate to form 2-methyl-6-geranylgeranylbenzoquinol. The HGGT gene is overexpressed using seeds-specific and constitutive promoters, leading to an increase of δ-TT con-tent of about two times around in the transgenic rice com-pared to the normal plant [147]. Similar enriched sources of TTs are obtained through the transformation of soybeans and Perilla frutescens, an annual plant native to Korean penin-sula, Southern China and India [148, 149].

3.3. Tocotrienols as Anti-Cancer Drugs

Recent patents suggest different methods for using TTs as anti-cancer drugs.

There is a recent invention according to which a diet of rice oil extract enriched in TTs can prevent colon cancer in F344 rats treated with the carcinogenic agent azoxymethane. In particular, the TTs treatment is reported to reduce the number and the volume of the aberrant crypt foci in the colonic mucosa of the animals, without causing any signifi-cant weight loss or side effect [150].

A new patent proposes γ-TT as a potent inhibitor of melanoma (C32 and G361), prostate cancer (LNCaP and PC-3) and breast cancer (MDA-MB-231 and MCF-7) cell growth both in vitro and in vivo. The anti-tumor effects of γ-TT were shown to be selective for cancer cells since normal cells, such as the immortalized human prostate epithelial PZ-HPV-7 cells, were not affected by the treatment, and they were associated with decreased cell proliferation and apopto-sis induction. In particular, γ-TT-mediated cytotoxicity was correlated with inhibition of IdI and NF-κB via modulation of their upstream regulators Src, Smadl/5/8, Fak and LOX. γ-TT treatment also resulted in the activation of JNK signaling pathway, and its inhibition by the specific inhibitor SP600125 partially restored cell viability. Interestingly, it was also found that γ-TT treatment enhanced the expression of E-cadherin, supporting the idea that γ-TT also possesses anti-metastatic activity in different tumors. Finally, it was demonstrated that γ-TT can down-regulate the expression of stem cell markers (CD133/CD44) [151].

A recent patent provides a method utilizing δ-TT to inhibit pancreatic tumor growth. Results show that δ-TT induces apoptosis and inhibits cell proliferation in human pancreatic ductal carcinoma cell lines (MIA PaCa2, SW1990, BXPC3)


in vitro as well as in MIA PaCa2 cells xenografted in nude mice. In particular, δ-TT induces caspases activation, decreases p-Akt, p-MAPK and Ki-67 levels and increases p27 levels. Immortalized human pancreatic ductal epithelial cells, HPDE 6C7, were treated under identical conditions, showing that δ-TT cytotoxic effects are selective for pancreatic cancer cells [152].

It should be noted that different inventions have evi-denced the cholesterol-lowering (due to HMG-CoA reduc-tase inactivation and mevalonate pathway inhibition) and the anti-oxidant (correlated with ROS suppression and block of superoxide release from tumor-associated neutrophils) prop-erties of TTs as well as their anti-angiogenic (consisting in the arrest of endothelial cell multiplication and lumen forma-tion) and immune-regulatory (associated with an increase in the total antibody titer) activity, suggesting the use of these compounds for cancer treatment [153-155].

3.4. Alternative Routes of Administration and Dosage Forms

As discussed above, TTs are usually administered per os. However, many studies have been focused on the devel-opment of new TTs formulations, and several patents have proposed alternative routes of administration and dosage forms for TTs delivery to target tissues.

There is a new patent that demonstrates that the transder-mal application of a gel containing δ-TT is very suitable and effective for the treatment of patients with breast cancer. Inter-estingly, δ-TT was observed to accumulate at higher concen-trations in the adipose tissue of the benign breast lumps in comparison to the malignant lumps, and this could be due to its antioxidant activity and in particular to its ability to quench free radicals and to regulate peroxidative reactions [156].

There is a recent invention according to which a nano-droplet containing a tocotrienol (e.g. δ-TT), a tocopherol or tocotrienol covalently bonded to a polyalkylene glycol (α-tocopheryl-polyethyleneglycol-1000-succinate), a poloxamer (PLURONIC® P188) and a polyalkylene glycol (PEG400) exerts a cytotoxic effect on NCI-H460 human lung cancer cells subcutaneously injected in nude mice. Moreover, it exhibits a synergistic anti-tumor effect in combination with docetaxel, and the severe weight loss caused by standard chemotherapy in the animals is reduced by simultaneous treatment with δ-TT nanodroplets, thus demonstrating their chemoprotective effects [157].

The Australian inventor Tong has found that the admini-stration of TTs by a transmucosal delivery route, particularly by sublingual tablets containing a dose of 9mg δ-TT and 1mg γ-TT, provides for enhanced bioavailability when com-pared with orally administered TTs, proposing it for cancer therapy [158]. Similarly, also the topical administration through the skin and the rectum has been suggested for TTs anti-cancer treatment [159].

3.5. Combinations with Other Compounds

Many inventors have tested one or more TTs in combi-nation with other compounds in different pathologies, in-cluding cancer.

A synergistic anti-tumor effect was observed when can-cer cells and xenografts were co-treated with γ-TT and a standard chemotherapeutic agent, such as docetaxel or

dacarbazine [151]. Similar results were also obtained after co-treatment with δ-TT and 5-fluorouracil (5-FU) in pancre-atic tumor cells [152].

A novel patent reports on a method for the prevention and treatment of neoplastic diseases and particularly of breast cancer, showing that a combination of citrus limon-oids (limonin or nomilin), citrus flavonoids (nobiletin or tangeretin) and TTs (α-, γ- or δ-TT), with or without conven-tional anti-tumor therapy (tamoxifen), synergistically inhibits MCF-7 and MDA-MB-231 cell proliferation [160].

Another invention for breast cancer treatment evidences that curcumin synergistically enhances the anti-proliferative effect of a TRF preparation on MCF-7 human breast cancer cells in vitro [161].

A recent patent provides a method using a composition comprising γ-TT and lovastatin or β-ionone to synergisti-cally inhibit the growth of B16 melanoma cells, also after their implantation in nude mice [162].

It has also been demonstrated that a combination of TTs with lovastatin and/or genistein can synergistically suppress prostate cancer (LNCaP, DU145 and PC-3) cell growth [163].

A new patent proposes the use of TTs gelatin capsules, to be administered orally and on a daily basis, for prostate and breast cancer treatment. Each capsule can contain 200mg of γ-TT, 75mg of δ-TT, 100mg of turmeric extract and 200mg of saw palmetto extract for prostate cancer therapy, and 100mg of γ-TT, 25mg of δ-TT, 100mg of turmeric extract and 50mg of L-ascorbic acid for breast cancer therapy. The composi-tional dosage can also include 100ml of fermented noni juice and 100ml of mangosteen juice, which can be administered orally separated from the gelatin capsules [164].

It has been recently suggested by an interesting patent that pharmaceutical compositions containing TTs and Met receptor tyrosine kinase inhibitors can exert a synergistic inhibitory effect on tumor cell growth. In cancer cells, Met can be consti-tutively activated via overexpression, mutations and/or inter-action with other membrane receptors, such as the HER/ErbB family members. Upregulation of the Met pathway can ulti-mately lead to increased tumor cell proliferation, reduced sus-ceptibility to apoptosis and enhanced angiogenesis and metas-tasis. Combinations of γ-TT and SU11274, a specific Met receptor tyrosine kinase inhibitor, synergistically inhibit the growth of +SA mammary tumor cells [165].

It has been shown by a novel patent that metallo-proteins, including lactoferrin, transferrin, ovotransferrin, ceruloplasmin and metallo-thionein, can stabilize and en-hance the bio-functional activity of TTs. Importantly, the synergism between metallo-proteins and TTs also promotes the intestinal transfer and the ultimate bioavailability of TTs for physiological functions [166].

Finally, different inventions present novel TTs antioxi-dant formulations that can be used to inhibit oxidative dam-age and to treat and prevent disorders caused by free radi-cals, including cancer. The antioxidant compositions com-prise: a combination of α-, β-, y- or δ-TT or TTs mixture plus a radical scavenger recycler (α-lipoic acid, ascorbic acid, ascorbic acid salts or ascorbyl palmitate) and optionally a radical formation inhibitor (pyruvate) [167]; a combination of δ-TT with green tea extract or pure epicatechin [168]; a combination of asthaxantin, betacarotene, lutein and a mix-


ture of TTs (56% of γ-TT, 30% of α-TT, 13% of δ-TT and 1% of other TTs including β-TT) [169].

3.6. Synthetic Derivatives

The first model of anti-cancer tocotrienol synthetic de-rivative was provided by Kanazawa et al., who proposed the linoleic acid α-TT ester as a preventive agent for cancer me-tastasis. Since then, several methods have been developed for the synthesis and use of TT derivatives [170].

There is a recent patent that indicates γ-TT succinate as a potent inhibitor of cancer cell growth, successfully suppress-ing the proliferation of liver cancer (Hep 3B), lung cancer (NIH-H69), colon cancer (HT29), prostate cancer (DU145), cutaneous melanoma (A375), ocular melanoma (OCMI), leu-kemia (HL60) and breast cancer (MCF7 and MD-231) cells [171].

A new patent reports that different ester derivatives of TTs exert both anti-proliferative and anti-invasive activity in +SA, MCF7 and MDA-MB-231 cell lines [172, 173].

A recent invention provides a method for synthetizing and using TTs ether derivatives. The novel compounds were shown to trigger apoptotic cell death of human breast cancer (MDA-MB-435, MDA-MB-231 and MCF-7), prostate can-cer (PC-3, DU145 and LnCaP), ovarian tumor (C-170), cer-vical tumor (ME-180), endometrial carcinoma (RL-95-2), leukemia (HL-60), colon cancer (HT-29 and DLD-1) and lung cancer (A-549) cells via activation of the TGF-β, stress-activated protein kinase and Fas/FasL signaling pathways. Moreover, they did not induce apoptosis of Normal Human Mammary Epithelial Cells (HMECs) and of immortalized but non-tumorigenic MCF-10A [174] mammary cells Table 2 [20, 128-174].

Table 2. Patents for the Isolation, Chemical Modification and Use of Tocotrienols (TTs) in Cancer Prevention/Therapy.

Patent Type Methods References

Isolation and

purification

• From red palm oil: 1) extraction from plant fatty acid distillates, conversion into distilled methyl esters, ion-exchange chromatography, molecular distillation; 2) conversion of crude palm oil to methyl esters, distilla-tion or chromatographic separation; 3) adsorption/desorption chromatography from crude palm oil with su-percritical fluids (i.e. SC-CO2)

• From rice bran: ultrasound-assisted saponification and recrystallization or chromatography separation • From coconut palm oil: transesterification, distillation and adsorption treatment with anion exchange resin • From TTs rich oils or distillates: flash or low-pressure chromatography

[20, 128-145]

Transgenic sources Rice plant, soybeans and Perilla Frutescens transformed through introduction of the Homogentisic Acid Geranylgeranyl Transferase (HGGT) gene [146-149]

TTs as anti-cancer drugs

• Anti-proliferative activity • Pro-apoptotic activity • Anti-metastatic activity • Anti-angiogenic activity • Cancer stem cells-targeting activity • Cholesterol-lowering activity • Anti-oxidant activity • Immune-regulatory activity

[150-155]

Alternative routes of administration and

dosage forms

• Transdermal gel

• Subcutaneous nanodroplets

• Sublingual tablets

[156-159]

Combinations

• Docetaxel or dacarbazine • Fluorouracil • Citrus limonoids and flavonoids • Curcumin • Lovastatin or β-ionone • Lovastatin and/or genistein • Turmeric extract and palmetto extract or L-ascorbic acid • Turmeric extract, palmetto extract, fermented noni juice and mangosteen juice • Met receptor tyrosine kinase inhibitors • Metallo-proteins: lactoferrin, transferrin, ovotransferrin, ceruloplasmin and metallothionein • α-Lipoic acid, ascorbic acid, ascorbic acid salts, ascorbyl palmitate and pyruvate • Green tea extract or pure epicatechin • Asthaxantin, betacarotene and lutein

[151, 152, 160-169]

Synthetic derivatives

• Linoleic acid α-TT ester • γ-TT succinate • Ester derivatives • Ether derivatives

[170-174]


CONCLUSION

The present work provides a review of recent literature and patents on the anti-cancer activity of TTs.

Several studies were conducted to investigate the effects of TTs on cancer growth and progression and to clarify the molecular mechanisms underlying these effects.

More recent studies focus on the development of new TTs synthetic derivatives, formulations and combinations, with the aim to open new windows of novel chemopreven-tive/therapeutic interventions.

The translation of these findings into the clinical setting still represents a great challenge and requires further investigation.

CURRENT & FUTURE DEVELOPMENTS

In recent years, interest in vitamin E-derived TTs has considerably increased owing to their unique health benefits and non-toxic nature.

Many in vitro and in vivo experiments have demon-strated the anti-tumor effects of TTs (especially γ-‐ and δ-‐TT) in several cancer cell lines and mouse models through the inhibition of cell proliferation and induction of apoptosis. Moreover, TTs have been shown to possess important anti-angiogenic and anti-metastatic properties and to specifically target and eliminate cancer stem cells.

The ability to act through multiple molecular pathways, such as by HMG-‐CoA reductase downregulation, growth factor receptors signaling cascades inhibition and ER stress and autophagy induction, makes these compounds ideal for sensitizing cancer cells to standard chemotherapeutic agents and to other anti-cancer natural compounds, as evidenced by different preclinical trials.

Based on these promising results, various inventions have been focused on the biotechnological development of novel TTs transgenic sources with the aim to enhance the natural availability of these compounds, and many studies have been conducted to design and synthesize new deriva-tives and formulations of TTs with the purpose to improve their biological activity.

Despite all these encouraging observations, the clinical data so far available are still incomplete and several impor-tant questions remain to be addressed about the role of TTs in cancer patients, especially as regards their pharmacokinet-ics and bioavailability. Thus, clinical trials aimed to clarify the TTs anti-tumor effectiveness are urgently needed.

ETHICS APPROVAL AND CONSENT TO PARTICI-PATE

Not applicable.

HUMAN AND ANIMAL RIGHTS

No Animals/Humans were used for studies that are the basis of this research.

CONSENT FOR PUBLICATION

Not applicable.

CONFLICT OF INTEREST

The authors declare no conflict of interest, financial or otherwise.

ACKNOWLEDGEMENTS

The authors would like to thank Comitato Emme Rouge Onlus for supporting Dr. Monica Marzagalli.

REFERENCES [1] Kalra EK. Nutraceutical: Definition and introduction. AAPS

PharmSci 2003; 5: E25. [2] Ahsan H, Ahad A, Iqbal J, Siddiqui WA. Pharmacological potential

of tocotrienols: A review. Nutr Metab (Lond) 2014; 11: 52-9. [3] Montagnani M.M, Marzagalli M, Fontana F, Raimondi M, Moretti

RM, Limonta P. Anticancer properties of tocotrienols: A review of cellular mechanisms and molecular targets. J Cell Physiol 2018: doi: 10.1002/jcp.27075.

[4] De Silva L, Chuah LH, Meganathan P, Fu JY. Tocotrienol and cancer metastasis. BioFactors 2016; 42: 149-62.

[5] Kannappan R, Gupta SC, Kim JH, Aggarwal BB. Tocotrienols fight cancer by targeting multiple cell signaling pathways. Genes Nutr 2012; 7: 43-52.

[6] Evans HM, Bishop KS. Existence of a hitherto unknown dietary factor essential for reproduction. J Am Med Assoc 1923; 81: 889-92.

[7] Evans HM. The isolation from wheat germ oil of an alcohol, α-tocopherol, having the properties of vitamin E. J Biol Chem 1936; 113: 319-32.

[8] Shibata A, Nakagawa K, Sookwong P, Tsuduki T, Asai A, Miyazawa T. α-Tocopherol attenuates the cytotoxic effect of δ-tocotrienol in human colorectal adenocarcinoma cells. Biochem Biophys Res Commun 2010; 397: 214-9.

[9] Uchida T, Abe C, Nomura S, Ichikawa T, Ikeda S. Tissue distribution of α- and γ-tocotrienol and γ-tocopherol in rats and interference with their accumulation by α-tocopherol. Lipids 2012; 47: 129-39.

[10] Khan MR, Siddiqui S, Parveen K, Javed S, Diwakar S, Siddiqui WA. Nephroprotective action of tocotrienol-rich fraction (TRF) from palm oil against potassium dichromate (K2Cr2O7)-induced acute renal injury in rats. Chem Biol Interact 2010; 186: 228-38.

[11] Khanna S, Patel V, Rink C, Roy S, Sen CK. Delivery of orally supplemented α-tocotrienol to vital organs of rats and tocopherol-transport protein deficient mice. Free Radic Biol Med 2005; 39: 1310-9.

[12] Qureshi AA, Khan DA, Mushtaq S, Ye SQ, Xiong M, Qureshi N. δ-Tocotrienol feeding modulates gene expression of EIF2, mTOR, protein ubiquitination through multiple-signaling pathways in chronic hepatitis C patients. Lipids Health Dis 2018; 17: 167.

[13] Bartosińska E, Jacyna J, Borsuk DA, Kaliszan M, Kruszewski WJ, Jankowski Z, et al. HPLC-APCI-MS/MS method development and validation for determination of tocotrienols in human breast adipose tissue. Talanta 2018; 176: 108-15.

[14] Springett GM, Husain K, Neuger A, Centeno B, Chen D-T, Hutchinson TZ, et al. A Phase I safety, pharmacokinetic, and pharmacodynamic presurgical trial of vitamin e δ-tocotrienol in patients with pancreatic ductal neoplasia. BioMedicine 2015; 2: 1987-95.

[15] Mei HN, Yuen MC, Ah NM, Cheng HC, Hashim MA. Separation of vitamin E (tocopherol, tocotrienol, and tocomonoenol) in palm oil. Lipids 2004; 39: 1031-5.

[16] Raddatz-Mota D, Pérez-Flores LJ, Carrari F, Mendoza-Espinoza JA, de León-Sánchez FD, Pinzón-López LL, et al. Achiote (Bixa orellana L.): A natural source of pigment and vitamin E. J Food Sci Technol 2017; 54: 1729-41.

[17] Goufo P, Trindade H. Rice antioxidants: Phenolic acids, flavonoids, anthocyanins, proanthocyanidins, tocopherols, tocotrienols, γ-oryzanol, and phytic acid. Food Sci Nutr 2014; 2: 75-104.

[18] Min B, McClung AM, Chen M-H. Phytochemicals and antioxidant capacities in rice brans of different color. J Food Sci 2011; 76: C117-26.


[19] Shahidi F, De Camargo AC. Tocopherols and tocotrienols in common and emerging dietary sources: Occurrence, applications, and health benefits. Int J Mol Sci 2016; 10: 1745.

[20] Ahsan H, Ahad A, Siddiqui WA. A review of characterization of tocotrienols from plant oils and foods. J Chem Biol 2015; 8: 45-59.

[21] Prasad K. Tocotrienols and cardiovascular health. Curr Pharm Des 2011; 17: 2147-54.

[22] Chin KY, Tay SS. A Review on the relationship between tocotrienol and Alzheimer disease. Nutrients 2018; 10: 881.

[23] Abdul-Majeed S, Mohamed N, Soelaiman I-N. A review on the use of statins and tocotrienols, individually or in combination for the treatment of osteoporosis. Curr Drug Targets 2013; 14: 1579-90.

[24] Shen CL, Klein A, Chin KY, Mo H, Tsai P, Yang RS, et al. Tocotrienols for bone health: A translational approach. Ann NY Acad Sci 2017; 1401: 150-65.

[25] Hsieh TC, Elangovan S, Wu JM. γ-Tocotrienol controls proliferation, modulates expression of cell cycle regulatory proteins and up-regulates quinone reductase NQO2 in MCF-7 breast cancer cells. Anticancer Res 2010; 30: 2869-74.

[26] Samant GV, Wali VB, Sylvester PW. Anti-proliferative effects of gamma-tocotrienol on mammary tumour cells are associated with suppression of cell cycle progression. Cell Prolif 2010; 43: 77-83.

[27] Parajuli P, Tiwari RV, Sylvester PW. Anti-proliferative effects of γ-tocotrienol are associated with suppression of c-Myc expression in mammary tumour cells. Cell Prolif 2015; 48: 421-35.

[28] Takahashi K, Loo G. Disruption of mitochondria during tocotrienol-induced apoptosis in MDA-MB-231 human breast cancer cells. Biochem Pharmacol 2004; 67: 315-24.

[29] Loganathan R, Selvaduray KR, Nesaretnam K, Radhakrishnan AK. Tocotrienols promote apoptosis in human breast cancer cells by inducing poly(ADP-ribose) polymerase cleavage and inhibiting nuclear factor kappa-B activity. Cell Prolif 2013; 46: 203-13.

[30] Shah S, Sylvester PW. Tocotrienol-induced caspase-8 activation is unrelated to death receptor apoptotic signaling in neoplastic mammary epithelial cells. Exp Biol Med 2004; 229: 745-755.

[31] Khallouki F, de Medina P, Caze-Subra S, Bystricky K, Balaguer P, Poirot M, et al. Molecular and biochemical analysis of the estrogenic and proliferative properties of vitamin E compounds. Front Oncol 2016; 5: 287.

[32] Viola V, Ciffolilli S, Legnaioli S, Piroddi M, Betti M, Mazzini F, et al. Mitochondrial-dependent anticancer activity of δ-tocotrienol and its synthetic derivatives in HER-2/neu overexpressing breast adenocarcinoma cells. BioFactors 2013; 39: 485-93.

[33] Asif HM, Sultana S, Ahmed S, Akhtar N, Tariq M. HER-2 positive breast cancer - A mini-review. Asian Pac J Cancer Prev 2016; 17: 1609-15.

[34] Alawin OA, Ahmed RA, Ibrahim BA, Briski KP, Sylvester PW. Antiproliferative effects of γ-tocotrienol are associated with lipid raft disruption in HER2-positive human breast cancer cells. J Nutr Biochem 2016; 27: 266-77.

[35] Comitato R, Leoni G, Canali R, Ambra R, Nesaretnam K, Virgili F. Tocotrienols activity in MCF-7 breast cancer cells: Involvement of ERβ signal transduction. Mol Nutr Food Res 2010; 54: 669-78.

[36] Iurlaro R, Muñoz-Pinedo C. Cell death induced by endoplasmic reticulum stress. FEBS J 2016; 283: 2640-52.

[37] Glick D, Barth S, Macleod KF. Autophagy: Cellular and molecular mechanisms. J Pathol 2010; 221: 3-12.

[38] Wang M, Law ME, Castellano RK, Law BK. The unfolded protein response as a target for anticancer therapeutics. Crit Rev Oncol Hematol 2018; 127: 66-79.

[39] Levy JMM, Towers CG, Thorburn A. Targeting autophagy in cancer. Nat Rev Cancer 2017; 17: 528-42.

[40] Kim C, Kim B. Anti-cancer natural products and their bioactive compounds inducing ER stress-mediated apoptosis: A review. Nutrients 2018; 10: 1021.

[41] Lin SR, Fu YS, Tsai MJ, Cheng H, Weng CF. Natural compounds from herbs that can potentially execute as autophagy inducers for cancer therapy. Int J Mol Sci 2017; 18: 1412-9.

[42] Wali VB, Bachawal SV, Sylvester PW. Endoplasmic reticulum stress mediates gamma-tocotrienol-induced apoptosis in mammary tumor cells. Apoptosis 2009; 14: 1366-77.

[43] Park SK, Sanders BG, Kline K. Tocotrienols induce apoptosis in breast cancer cell lines via an endoplasmic reticulum stress-dependent increase in extrinsic death receptor signaling. Breast Cancer Res Treat 2010; 124: 361-75.

[44] Patacsil D, Tran AT, Cho YS, Suy S, Saenz F, Malyukova I, et al. Gamma-tocotrienol induced apoptosis is associated with unfolded protein response in human breast cancer cells. J Nutr Biochem 2011; 23: 93-100.

[45] Tiwari RV, Parajuli P, Sylvester PW. γ-Tocotrienol-induced autophagy in malignant mammary cancer cells. Exp Biol Med 2014; 239: 33-44

[46] Tiwari RV, Parajuli P, Sylvester PW. γ-Tocotrienol-induced endoplasmic reticulum stress and autophagy act concurrently to promote breast cancer cell death. Biochem Cell Biol 2015; 93: 306-20.

[47] Sylvester PW. Targeting met mediated epithelial-mesenchymal transition in the treatment of breast cancer. Clin Transl Med 2014; 3: 30.

[48] Selvaduray KR, Radhakrishnan AK, Kutty MK, Nesaretnam K. Palm tocotrienols inhibit proliferation of murine mammary cancer cells and induce expression of interleukin-24 mRNA. J Interf Cytokine Res 2010; 30: 909-16.

[49] Selvaduray KR, Radhakrishnan AK, Kutty MK, Nesaretnam K. Palm tocotrienols decrease levels of pro-angiogenic markers in human umbilical vein endothelial cells (HUVEC) and murine mammary cancer cells. Genes Nutr 2012; 7: 53-61.

[50] Algayadh IG, Dronamraju V, Sylvester PW. Role of Rac1/WAVE2 signaling in mediating the inhibitory effects of gamma-tocotrienol on mammary cancer cell migration and invasion. Biol Pharm Bull 2016; 39: 1974-82.

[51] Nassar D, Blanpain C. Cancer stem cells: Basic concepts and therapeutic implications. Annu Rev Pathol Mech Dis 2016; 11: 47-76.

[52] Taylor WF, Jabbarzadeh E. The use of natural products to target cancer stem cells. Am J Cancer Res 2017; 7: 1588-605.

[53] Gopalan A, Yu W, Sanders BG, Kline K. Eliminating drug resistant breast cancer stem-like cells with combination of simvastatin and gamma-tocotrienol. Cancer Lett 2013; 328: 285-96.

[54] Xiong A, Yu W, Liu Y, Sanders BG, Kline K. Elimination of ALDH+ breast tumor initiating cells by docosahexanoic acid and/or gamma tocotrienol through SHP-1 inhibition of Stat3 signaling. Mol Carcinog 2016; 55: 420-30.

[55] Bachawal SV, Wali VB, Sylvester PW. Combined γ-tocotrienol and erlotinib/gefitinib treatment suppresses Stat and Akt signaling in murine mammary tumor cells. Anticancer Res 2010; 30: 429-37.

[56] Wali VB, Sylvester PW. Synergistic antiproliferative effects of γ-tocotrienol and statin treatment on mammary tumor cells. Lipids 2007; 42: 1113-23.

[57] B. Shirode A, W. Sylvester P. Mechanisms mediating the synergistic anticancer effects of combined γ-tocotrienol and celecoxib treatment. J Bioanal Biomed 2011; 3: 1-7.

[58] Ding Y, Peng Y, Deng L, Fan J, Huang B. Gamma-tocotrienol reverses multidrug resistance of breast cancer cells with a mechanism distinct from that of atorvastatin. J Steroid Biochem Mol Biol 2017; 167: 67-77.

[59] Sontag TJ, Parker RS. Cytochrome P450 omega-hydroxylase pathway of tocopherol catabolism. Novel mechanism of regulation of vitamin E status. J Biol Chem 2002; 277: 25290-6.

[60] Akl MR, Ayoub NM, Sylvester PW. Mechanisms mediating the synergistic anticancer effects of combined γ-tocotrienol and sesamin treatment. Planta Med 2012; 78: 1731-9.

[61] Akl MR, Ayoub NM, Abuasal BS, Kaddoumi A, Sylvester PW. Sesamin synergistically potentiates the anticancer effects of γ-tocotrienol in mammary cancer cell lines. Fitoterapia 2013; 84: 347-59.

[62] Hsieh TC, Wu JM. Suppression of cell proliferation and gene expression by combinatorial synergy of EGCG, resveratrol and γ-tocotrienol in estrogen receptor-positive MCF-7 breast cancer cells. Int J Oncol 2008; 33: 851-9

[63] Tiwari R V., Parajuli P, Sylvester PW. Synergistic anticancer effects of combined γ-tocotrienol and oridonin treatment is associated with the induction of autophagy. Mol Cell Biochem 2015; 408: 123-37.

[64] Behery FA, Akl MR, Ananthula S, Parajuli P, Sylvester PW, El Sayed KA. Optimization of tocotrienols as antiproliferative and antimigratory leads. Eur J Med Chem 2013; 59: 329-41.

[65] Ananthula S, Parajuli P, Behery FA, Alayoubi AY, El Sayed KA, Nazzal S, et al. Oxazine derivatives of γ- and δ-tocotrienol display enhanced anticancer activity in vivo. Anticancer Res 2014; 34: 2715-26.


[66] Ananthula S, Parajuli P, Behery FA, Alayoubi AY, Nazzal S, El Sayed K, et al. δ-Tocotrienol oxazine derivative antagonizes mammary tumor cell compensatory response to COCl2-induced hypoxia. Biomed Res Int 2014; 2014: 285752.

[67] Nesaretnam K, Selvaduray KR, Abdul Razak G, Veerasenan SD, Gomez PA. Effectiveness of tocotrienol-rich fraction combined with tamoxifen in the management of women with early breast cancer: a pilot clinical trial. Breast Cancer Res 2010; 12: R81.

[68] Wu S-J, Ng L-T. Tocotrienols inhibited growth and induced apoptosis in human HeLa cells through the cell cycle signaling pathway. Integr Cancer Ther 2010, 9: 66-72.

[69] Comitato R, Guantario B, Leoni G, Nesaretnam K, Ronci MB, Canali R, et al. Tocotrienols induce endoplasmic reticulum stress and apoptosis in cervical cancer cells. Genes Nutr 2016; 11: 32.

[70] Gu W, Prasadam I, Yu M, Zhang F, Ling P, Xiao Y, et al. Gamma tocotrienol targets tyrosine phosphatase SHP2 in mammospheres resulting in cell death through RAS/ERK pathway. BMC Cancer 2015; 15: 609.

[71] Agarwal MK, Agarwal ML, Athar M, Gupta S. Tocotrienol-rich fraction of palm oil activates p53, modulates Bax/Bcl2 ratio and induces apoptosis independent of cell cycle association. Cell Cycle 2004; 3: 205-11.

[72] Jang Y, Rao X, Jiang Q. Gamma-tocotrienol profoundly alters sphingolipids in cancer cells by inhibition of dihydroceramide desaturase and possibly activation of sphingolipid hydrolysis during prolonged treatment. J Nutr Biochem 2017; 46: 49-56.

[73] Lee D, Kim IY, Saha S, Choi KS. Paraptosis in the anti-cancer arsenal of natural products. Pharmacol Ther 2016; 162: 120-33.

[74] Zhang JS, Li DM, Ma Y, He N, Gu Q, Wang F-S, et al. γ-Tocotrienol induces paraptosis-like cell death in human colon carcinoma SW620 cells. PLoS One 2013; 8: e57779.

[75] Zhang JS, Li DM, He N, Liu YH, Wang CH, Jiang S-Q, et al. A paraptosis-like cell death induced by δ-tocotrienol in human colon carcinoma SW620 cells is associated with the suppression of the Wnt signaling pathway. Toxicology 2011; 285: 8-17.

[76] Prasad S, Gupta SC, Tyagi AK, Aggarwal BB. γ-Tocotrienol suppresses growth and sensitises human colorectal tumours to capecitabine in a nude mouse xenograft model by down-regulating multiple molecules. Br J Cancer 2016; 115: 814-824.

[77] Yang Z, Xiao H, Jin H, Koo PT, Tsang DJ, Yang CS. Synergistic actions of atorvastatin with gamma-tocotrienol and celecoxib against human colon cancer HT29 and HCT116 cells. Int J Cancer 2010; 126: 852-63.

[78] Md Yusof K, Makpol S, Jamal R, Harun R, Mokhtar N, Wan Ngah W. γ-Tocotrienol and 6-gingerol in combination synergistically induce cytotoxicity and apoptosis in HT-29 and SW837 human colorectal cancer cells. Molecules 2015; 20: 10280-97.

[79] Shibata A, Nakagawa K, Sookwong P, Tsuduki T, Tomita S, Shirakawa H, et al. Tocotrienol inhibits secretion of angiogenic factors from human colorectal adenocarcinoma cells by suppressing Hypoxia-Inducible Factor-1α. J Nutr 2008; 138: 2136-42.

[80] Shibata A, Nakagawa K, Sookwong P, Tsuzuki T, Oikawa S, Miyazawa T. Tumor anti-angiogenic effect and mechanism of action of δ-tocotrienol. Biochem Pharmacol 2008; 76: 330-9.

[81] Shibata A, Nakagawa K, Sookwong P, Tsuduki T, Oikawa S, Mlyazawa T. δ-tocotrienol suppresses VEGF induced angiogenesis whereas α-tocopherol does not. J Agric Food Chem 2009; 57: 8696-704.

[82] Eitsuka T, Tatewaki N, Nishida H, Nakagawa K, Miyazawa T. A combination of δ-tocotrienol and ferulic acid synergistically inhibits telomerase activity in DLD-1 human colorectal adenocarcinoma cells. J Nutr Sci Vitaminol 2016; 62: 281-287.

[83] Sun W, Wang Q, Chen B, Liu J, Liu H, Xu W. Gamma-tocotrienol-induced apoptosis in human gastric cancer SGC-7901 cells is associated with a suppression in mitogen-activated protein kinase signalling. Br J Nutr 2008; 99: 1247-54.

[84] Sun W, Xu W, Liu H, Liu J, Wang Q, Zhou J, et al. γ-Tocotrienol induces mitochondria-mediated apoptosis in human gastric adenocarcinoma SGC-7901 cells. J Nutr Biochem 2009; 20: 276-84.

[85] Liu H-K, Wang Q, Li Y, Sun W-G, Liu J-R, Yang Y-M, et al. Inhibitory effects of γ-tocotrienol on invasion and metastasis of human gastric adenocarcinoma SGC-7901 cells. J Nutr Biochem 2010; 21: 206-13.

[86] Bi S, Liu JR, Li Y, Wang Q, Liu HK, Yan YG, et al. γ-Tocotrienol modulates the paracrine secretion of VEGF induced by cobalt(II) chloride via ERK signaling pathway in gastric adenocarcinoma SGC-7901 cell line. Toxicology 2010; 274: 27-33.

[87] Li Y, Sun W-G, Liu H-K, Qi G-Y, Wang Q, Sun X-R, et al. γ-Tocotrienol inhibits angiogenesis of human umbilical vein endothelial cell induced by cancer cell. J Nutr Biochem 2011; 22: 1127-36.

[88] Manu KA, Shanmugam MK, Ramachandran L, Li F, Fong CW, Kumar AP, et al. First evidence that γ-tocotrienol inhibits the growth of human gastric cancer and chemosensitizes it to capecitabine in a xenograft mouse model through the modulation of NF-κB pathway. Clin Cancer Res 2012; 18: 2220-9.

[89] Peng Y, Croce CM. The role of microRNAS in human cancer. Signal Transduct Target Ther 2016; 1: 15004.

[90] Ji X, Wang Z, Geamanu A, Goja A, Sarkar FH, Gupta S V. Delta-tocotrienol suppresses Notch-1 pathway by upregulating miR-34a in nonsmall cell lung cancer cells. Int J Cancer 2012; 131: 2668-77.

[91] Ji X, Wang Z, Sarkar FH, Gupta S V. Delta-tocotrienol augments cisplatin-induced suppression of non-small cell lung cancer cells via inhibition of the Notch-1 pathway. Anticancer Res 2012; 32: 2647-55.

[92] McAnally J a, Gupta J, Sodhani S, Bravo L, Mo H. Tocotrienols potentiate lovastatin-mediated growth suppression in vitro and in vivo. Exp Biol Med (Maywood) 2007; 232: 523-31.

[93] Hodul PJ, Dong Y, Husain K, Pimiento JM, Chen J, Zhang A, et al. Vitamin E δ-Tocotrienol induces p27Kip1-dependent cell-cycle arrest in pancreatic cancer cells via an E2F-1-dependent mechanism. PLoS One 2013; 8: e52526.

[94] Hussein D, Mo H. d-δ-Tocotrienol-mediated suppression of the proliferation of human PANC-1, MIA PaCa-2, and BxPC-3 Pancreatic Carcinoma Cells. Pancreas 2009; 38: e124-36.

[95] Shin-Kang S, Ramsauer VP, Lightner J, Chakraborty K, Stone W, Campbell S, et al. Tocotrienols inhibit AKT and ERK activation and suppress pancreatic cancer cell proliferation by suppressing the ErbB2 pathway. Free Radic Biol Med 2011; 51: 1164-1174.

[96] Wang C, Husain K, Zhang A, Centeno BA, Chen D-T, Tong Z, et al. EGR-1/Bax pathway plays a role in vitamin E δ-tocotrienol-induced apoptosis in pancreatic cancer cells. J Nutr Biochem 2015; 26: 797-807.

[97] Kunnumakkara AB, Sung B, Ravindran J, Diagaradjane P, Deorukhkar A, Dey S, et al. γ-tocotrienol inhibits pancreatic tumors and sensitizes them to gemcitabine treatment by modulating the inflammatory microenvironment. Cancer Res 2010; 70: 8695-705.

[98] Husain K, Centeno BA, Coppola D, Trevino J, Sebti SM, Malafa MP. δ-Tocotrienol, a natural form of vitamin E, inhibits pancreatic cancer stem-like cells and prevents pancreatic cancer metastasis. Oncotarget 2017; 8: 31554-31567.

[99] Abu-Fayyad A, Nazzal S. Synthesis, characterization, and in vitro antitumor activity of the polyethylene glycol (350 and 1000) succinate derivatives of the tocopherol and tocotrienol isomers of Vitamin E. Int J Pharm 2017; 519: 145-56.

[100] Abu-Fayyad A, Nazzal S. Gemcitabine-vitamin E conjugates: Synthesis, characterization, entrapment into nanoemulsions, and in-vitro deamination and antitumor activity. Int J Pharm 2017; 528: 463-70.

[101] Abu-Fayyad A, Kamal MM, Carroll JL, Dragoi A-M, Cody R, Cardelli J, et al. Development and in vitro characterization of nanoemulsions loaded with paclitaxel/γ-tocotrienol lipid conjugates. Int J Pharm 2018; 536: 146-57.

[102] Yap WN, Chang PN, Han HY, Lee DTW, Ling MT, Wong YC, et al. Gamma-tocotrienol suppresses prostate cancer cell proliferation and invasion through multiple-signalling pathways. Br J Cancer 2008; 99: 1832-41.

[103] Barve A, Khor TO, Reuhl K, Reddy B, Newmark H, Kong A-N. Mixed tocotrienols inhibit prostate carcinogenesis in TRAMP mice. Nutr Cancer 2010; 62: 789-94.

[104] Campbell SE, Rudder B, Phillips RB, Whaley SG, Stimmel JB, Leesnitzer LM, et al. γ-Tocotrienol induces growth arrest through a novel pathway with TGFβ2 in prostate cancer. Free Radic Biol Med 2011: 50: 1344-54.

[105] Sugahara R, Sato A, Uchida A, Shiozawa S, Sato C, Virgona N, et al. Annatto tocotrienol induces a cytotoxic effect on human prostate cancer PC3 cells via the simultaneous inhibition of Src and Stat3. J Nutr Sci Vitaminol (Tokyo) 2015; 61: 497-501.


[106] Huang Y, Wu R, Su ZY, Guo Y, Zheng X, Yang CS, et al. A naturally occurring mixture of tocotrienols inhibits the growth of human prostate tumor, associated with epigenetic modifications of cyclin-dependent kinase inhibitors p21 and p27. J Nutr Biochem 2017; 40: 155-63.

[107] Luk SU, Yap WN, Chiu YT, Lee DTW, Ma S, Lee TKW, et al. Gamma-tocotrienol as an effective agent in targeting prostate cancer stem cell-like population. Int J Cancer 2011; 128: 2182-91.

[108] Lee SO, Ma Z, Yeh C-R, Luo J, Lin T-H, Lai K-P, et al. New therapy targeting differential androgen receptor signaling in prostate cancer stem/progenitor vs. non-stem/progenitor cells. J Mol Cell Biol 2013; 5: 14-26.

[109] Kaneko S, Sato C, Shiozawa N, Sato A, Sato H, Virgona N, Yano T. Suppressive Effect of delta-tocotrienol on hypoxia adaptation of prostate cancer stem-like cells. Anticancer Res 2018; 38(3): 1391-9.

[110] Mo H, Elson CE. Studies of the isoprenoid-mediated inhibition of mevalonate synthesis applied to cancer chemotherapy and chemoprevention. Exp Biol Med (Maywood) 2004; 229: 567-85.

[111] Yeganehjoo H, DeBose-Boyd R, McFarlin BK, Mo H. Synergistic impact of d-δ-tocotrienol and geranylgeraniol on the growth and HMG CoA reductase of human DU145 Prostate carcinoma cells. Nutr Cancer 2017; 69: 682-91.

[112] Montagnani Marelli M, Marzagalli M, Moretti RM, Beretta G, Casati L, Comitato R, et al. Vitamin E δ-tocotrienol triggers endoplasmic reticulum stress-mediated apoptosis in human melanoma cells. Sci Rep 2016; 6: 30502.

[113] Marzagalli M, Moretti RM, Messi E, Montagnani Marelli M, Fontana F, Anastasia A, et al. Targeting melanoma stem cells with the Vitamin E derivative δ-tocotrienol. Sci Rep 2018; 8: 587.

[114] Chang PN, Yap WN, Lee DTW, Ling MT, Wong YC, Yap YL. Evidence of gamma-tocotrienol as an apoptosis-inducing, invasion-suppressing, and chemotherapy drug-sensitizing agent in human melanoma cells. Nutr Cancer 2009; 61: 357-66.

[115] Kawajiri K, Fujii-Kuriyama Y. The aryl hydrocarbon receptor: A multifunctional chemical sensor for host defense and homeostatic maintenance. Exp Anim 2017; 66: 75-89.

[116] Yamashita S, Baba K, Makio A, Kumazoe M, Huang Y, Lin I-C, et al. γ-Tocotrienol upregulates aryl hydrocarbon receptor expression and enhances the anticancer effect of baicalein. Biochem Biophys Res Commun 2016; 473: 801-7.

[117] Pham J, Nayel A, Hoang C, Elbayoumi T. Enhanced effectiveness of tocotrienol-based nano-emulsified system for topical delivery against skin carcinomas. Drug Deliv 2016; 23: 1514-24.

[118] Ledet G, Biswas S, Kumar V, Graves R, Mitchner D, Parker T, et al. Development of orally administered γ-tocotrienol (GT3) nanoemulsion for radioprotection. Int J Mol Sci 2016; 18: 28.

[119] Karim R, Somani S, Al Robaian M, Mullin M, Amor R, McConnell G, et al. Tumor regression after intravenous administration of targeted vesicles entrapping the vitamin E α-tocotrienol. J Control Release 2017; 246: 79-87.

[120] Ye C, Zhao W, Li M, Zhuang J, Yan X, Lu Q, et al. δ-Tocotrienol induces human bladder cancer cell growth arrest, apoptosis and chemosensitization through inhibition of STAT3 pathway. PLoS One 2015; 10: e0122712.

[121] Tan JK, Then SM, Mazlan M, Raja Abdul Rahman RNZ, Jamal R, Wan Ngah WZ. Gamma-tocotrienol acts as a BH3 mimetic to induce apoptosis in neuroblastoma SH-SY5Y cells. J Nutr Biochem 2016; 31: 28-37.

[122] Inoue A, Takitani K, Koh M, Kawakami C, Kuno T, Tamai H. Induction of apoptosis by α-Tocotrienol in human cancer cell lines and leukemic blasts from patients: Dependency on bid, cytochrome c, and caspase pathway. Nutr Cancer 2011; 63: 763-770.

[123] Wilankar C, Khan NM, Checker R, Sharma D, Patwardhan R, Gota V, et al. γ-Tocotrienol induces apoptosis in human T cell lymphoma through activation of both intrinsic and extrinsic pathways. Curr Pharm Des 2011; 17: 2176-89.

[124] Burdeos GC, Ito J, Eitsuka T, Nakagawa K, Kimura F, Miyazawa T. δ and γ tocotrienols suppress human hepatocellular carcinoma cell proliferation: Via regulation of Ras-Raf-MEK-ERK pathway-associated upstream signaling. Food Funct 2016; 7: 4170-74.

[125] Siveen KS, Ahn KS, Ong TH, Shanmugam MK, Li F, Yap WN, Kumar AP, Fong CW, Tergaonkar V, Hui KM, Sethi G. Gamma-tocotrienol inhibits angiogenesis-dependent growth of human hepatocellular carcinoma through abrogation of AKT/mTOR

pathway in an orthotopic mouse model. Oncotarget 2014;5(7):1897-911.

[126] Fu JY, Htar TT, De Silva L, Tan DMY, Chuah LH. Chromatographic Separation of vitamin E enantiomers. Molecules 2017; 22: 233.

[127] Beretta G, Gelmini F, Fontana F, Moretti RM, Montagnani Marelli M, Limonta P. Semi-preparative HPLC purification of δ-tocotrienol (δ-T3) from Elaeis guineensis Jacq. and Bixa orellana L. and evaluation of its in vitro anticancer activity in human A375 melanoma cells. Nat Prod Res 2018; 32: 1130-5.

[128] Top, A.G., Leong, L.W., Ong, A.S.H., Kawada, T., Watanabe, H., Tsuchiya, N. Production of high concentration tocopherols and tocotrienols from palm-oil by-products. US5190618 (1993).

[129] Furuse, T., Iwama, T., Kishima, S., Sakaguchi, T., Ueishi, S. Method for producing fatty acid concentrate of tocopherol and tocotrienol. JP2002003488 (2002).

[130] Ichitani, T., Tanaka, Y. Tocopherol concentrate and tocotrienol concentrate, and method for producing the same. JP2003171376 (2003).

[131] Kishima, S., Nakatate, M. Method for producing tocopherol and tocotrienol. JP2004305155 (2004).

[132] Jacobs, L. Process for the production of tocotrienols. US6838104 (2005).

[133] Dai, Z., Li, X., Zhao, J., Wang, S., Xu, X., Shao, B. Preparation method of high-purity tocopherol and tocotrienol concentrate mixture. CN106632209 (2017).

[134] Yin, J., Zhang, Y., Huang, Y., Huang, H. High-efficiency tocotrienol purifying method. CN107417658 (2017).

[135] Tan, B., Saleh, M.H. Integrated process for recovery of carotenoids and tocotrienols from oil. US5157132 (1992).

[136] Tou, G.P. Recovery of minor components from vegetable oils and fats. GB2387390 (2003).

[137] Shaohua, L., Nana, M., Caiyun, M., Shije, Z., Jingfang, Z. Natural carotenoid and tocotrienol-enriched red palm oil. CN102187914 (2011).

[138] Goh, R.S.H., Kam, R.T.S. Recovery of carotenoids, tocopherols, tocotrienols and sterols from esterified palm oil. GB2218989 (1991).

[139] Choo, Y.M., Ma, A.N., Basiron, D.Y. A method of chromatographic isolation for vitamin E isomers. CA2334068 (2010).

[140] Zhao, Y., Sun, M., Huang, S., Sun, R. Method for enriching mixture of natural totaxin and natural tocotrienol and extraction system. CN106215448 (2018).

[141] Lee, Y.S., Park, S.R., Kim, Y.H., Kang C.S. A method of mass-production tocotrienol in rice bran and bran-based functional foodstuff containing tocotrienol with high content. KR100884570 (2009).

[142] Maeda, Y., Hoang, T.T., Takahashi, H., Matsubara, T. Method for producing bioactive substance from rice bran. JP6335026 (2018).

[143] Tan, B, Foley, J. Tocotrienols and geranylgeraniol from Bixa orellana byproducts. US6350453 (2002).

[144] Honma, R., Abe, H., Ishikura, Y. Method for producing A tocotrienol composition. JP5272073 (2013).

[145] Howard, L. Process of producing purified gamma- and delta-tocotrienols from tocol-rich oils or distillates. US9512098 (2016).

[146] Babura SR, Abdullah SNA, Khaza A.H. Advances in genetic improvement for tocotrienol production: A review. J Nutr Sci Vitaminol (Tokyo) 2017; 63: 215-21.

[147] Park, H.M., Kim, Y.H., Lee, Y.Y., Lee, S.B., Lee, J.S. Composition for preventing or treating prostate cancer comprising rice transformed with tocotrienol biosynthesis gene. KR101629370 (2016).

[148] Kim, Y.H., Lee, Y.Y., Park, H.M., Choi, M.S., Yun, H.T., Lee, C.K., Kim W.H. Germinated transformants synthesizing tocotrienol with high anti-oxidant activity. KR101303741 (2015).

[149] Lee, Y.S., Tariq, H.M.M.H., Kim, K.H., Kim, Y.H. Transgenic perilla having increased tocotrienol content. KR100970379 (2010).

[150] Dobashi, K., Mori, H., Murai, H., Okada, T., Yoshimi, N. Colon cancer-preventing agent. JP2002316940 (2002).

[151] Ling, M.T., Yap, W.N., Wong, Y.C., Yap, Y.L.D. Use of tocotrienol composition for the prevention of cancer. US20110195910 (2011).

[152] Malafa, M.P., Sebti, S.M. Delta-tocotrienol treatment and prevention of pancreatic cancer. US8288369 (2012).


[153] Lane, R.H., Qureshi, A.A., Salser, W.A. Tocotrienols and tocotrienol-like compounds and methods for their use. US5591772 (1997).

[154] Ikushima, H., Miyazawa, T. Tocotrienols as inhibitors for neovascularization. EP1230923 (2006).

[155] Yamada, K., Yoshitake, S., Asano, T. Immune function improving agent. JP4271742 (2009).

[156] Nesadurai, K., Selvaduray, K.R., Hafid, S.R.A., Razak G.A., Huat O.T. Transdermal fluid. US20080194677 (2008).

[157] Tien, Y.D. Chemoprotective/chemoactive nanodroplets and methods of use thereof. WO2018005412 (2018).

[158] Tong, G. Transmucosal delivery of tocotrienol. WO2014075135 (2014).

[159] Erfinder, W.S.G. Use of an active ingredient consisting of e.g. tocotrienols in the substantial absence of tocopherols, for the prevention or treatment of e.g. normal or pathological inflammatory diseases, pain, diseases of nervous system, and cancer. DE102012020542 (2014).

[160] Guthrie, N., Kurowska, E.M., Carroll, K.K. Compositions and methods of treatment of neoplastic diseases and hypercholesterolemia with citrus limonoids and flavonoids and tocotrienols. US6251400 (2001).

[161] Nesaretnam, K., Selvaduray, K.R. Synergistic effect of tocotrienols and curcumin. US8906960 (2014).

[162] Elson, C.E. Method of suppressing tumor growth with combinations of isoprenoids and statins. EP0986383 (2000).

[163] Mo, H., Elson, C.E., Peffley, D.M., Hentosh, P.M. Composition and method for treating cancer. US7074825 (2006).

[164] Bellafiore, L., Radosevich, J.A., Glicken, E. Tocotrienol compositions. US20110293753 (2011).

[165] Sylvester, P.W. Anticancer combination treatments. US20130143866 (2013).

[166] Naidu, A.S., Naidu, A.G.S., Naidu, A.G.T. Metallo-protein and tocotrienol (MP-T3) compositions and uses thereof. US8309080 (2012).

[167] Schneider, F.H., Lane, R.H., Avila, T. Novel antioxidant formulations and methods for using them. WO2000057876 (2000).

[168] Hasler-Nguyen, N., Zijlstra, J., Troup, J.P. Synergistic antioxidant combination of delta tocols and polyphenols. US7452549 (2008).

[169] Babish, J., Howell, T. Compositions containing carotenoids and tocotrienols and having synergistic antioxidant effect. US20030206972 (2003).

[170] Kanazawa, A., Unnaka, Y., Wakabayashi, T. Unsaturated fatty acid derivative. JPS59222487 (1984).

[171] Fariss, M., Smith, J.D. Anti-tumor activity of vitamin E, cholesterol, taxol and betulinic acid derivatives. EP1189607 (2002).

[172] Sylvester, P.W., El Sayed, K.A. Anti-cancer tocotrienol analogues and associated methods. US8268786 (2012).

[173] El Sayed, K.A., Sylvester, P.W. Tocotrienol esters. US8969303 (2015).

[174] Gardner, R., Hurley, L., Israel, K., Kline, K., Liu, S., Menchaca, M., Ramanan, P.N., Sanders, B.G., Yu W. Tocopherols, tocotrienols, other chroman and side chain derivatives and uses thereof. EP1115398 (2010).

recent patents on anti-cancer drug discovery, 2019, 14, 5-18 · tts are wheat germ, grapefruit,...

Documents